显微成像分析技术在中性粒细胞运动及吞噬功能研究中的应用  

Applications of Microscopic Imaging and Analysis Technology in Studies of Neutrophil Movement and Phagocytosis

作　　者：杨晚竹[1] 高亚男[1] 梁昊岳 程雪莲[1] 于文颖[1] 陈婷[1] 汪晓敏[1] 袁卫平[1] 

机构地区：[1]中国医学科学院北京协和医学院血液病医院血液学研究所实验血液学国家重点实验室,天津300020

出　　处：《中国实验血液学杂志》2015年第3期832-837,共6页Journal of Experimental Hematology

基　　金：国家自然基金(81300436;81470280);实验血液学国家重点实验室自由申请项目(ZZ13-05)

摘　　要：目的:使用转盘式激光共聚焦显微镜及显微图像处理软件对中性粒细胞的运动及吞噬功能进行分析与评估。方法:使用Percoll分离液分离小鼠骨髓中性粒细胞,用PE荧光标记后与FITC标记的酵母聚糖颗粒(Zymosan)混匀,置于转盘式激光共聚焦显微镜下进行多通道时间序列成像。使用Volocity和Image J分析中性粒细胞的运动及吞噬相关参数,包括形态变化、运动轨迹、伪足振荡、结合吞噬指数,得出时间序列下各项参数的变化趋势。结果:中性粒细胞在受到Zymosan诱导后趋化作用明显,大部分细胞在短时间内发生极性化,在40分钟内能快速结合Zymosan,行使吞噬功能。结论:使用显微成像技术结合图像处理分析技术可准确、定量地分析中性粒细胞运动及吞噬功能,直观快速地对中性粒细胞的生理学状态进行初步评估。通过这项技术,得到正常生理状态下中性粒细胞对Zymosan的快速响应情况,为今后中性粒细胞相关功能学研究提供了参考依据。Objective： To analyze and evaluate the application of spinning disk confocal microscopy and imaging analysis software in movement and phagocytosis of neutrophils. Methods ： Neutrophils were isolated from bone marrow by centrifugation on discontinuous Percoll gradient, and then were stained with PE Gr-1 antibody and mixed with FITClabeled Zymosan A bioparticles. Multichannel time-lapse videos were captured by using the spinning disk confocal microscopy. The result was analyzed by using volocity and ImageJ software, the parameters associated with movement and phagocytosis of neutrophils were analyzed, including morphological changes, cell tracking, pseudopod dynamics, binding and phagocytosis index. Results： Most neutrophils would be polarized in response to Zymosan particles during a short time. Binding and phagocytosis process occured in forty minutes. Conclusion： A method of precisely quantifying the movement and phagocytosis of neutrophils using microscopic imaging and imaging analysis technique has been set up successfully. Using this method, biological activity and function of neutrophils can be evaluated visually and rapidly. The physiologically rapid response to Zymosan particles can be applied to the neutrophils function research in the future.

关 键 词：显微成像技术 图像定量分析 中性粒细胞 

分 类 号：R445[医药卫生—影像医学与核医学] R331.142[医药卫生—诊断学]